Dihydrosphingosine-Induced Programmed Cell Death in Tobacco BY-2 Cells Is Independent of H2O2 Production

Abstract

Sphinganine or dihydrosphingosine (d18:0, DHS), one of the most abundant free sphingoid Long Chain Base (LCB) in plants, has been recently shown to induce both cytosolic and nuclear calcium transient increases and a correlated Programmed Cell Death (PCD) in tobacco BY-2 cells. In this study, in order to get deeper insight into the LCB signaling pathway leading to cell death, the putative role of Reactive Oxygen Species (ROS) has been investigated. We show that DHS triggers a rapid dose-dependent production of H₂O₂ that is blocked by diphenyleniodonium (DPI), indicating the involvement of NADPH oxidase(s) in the process. In addition, while DPI does not block DHS-induced calcium increases, the ROS production is inhibited by the broad spectrum calcium channel blocker lanthanum (La³+). Therefore, ROS production occurs downstream of DHS-induced Ca²+ transients. Interestingly, DHS activates expression of defense-related genes that is inhibited by both La³+ and DPI. Since DPI does not prevent DHS-induced cell death, these results strongly indicate that DHS-induced H₂O₂ production is not implicated in PCD mechanisms but rather would be associated to basal cell defense mechanisms.

Nitric oxide production is not required for dihydrosphingosine-induced cell death in tobacco BY-2 cells

“…As previously shown, 8,10 external application of DHSinduced PCD in tobacco cells. The cell death ratio reached approximately 70% after a one hour treatment with 25 μM DHS (Fig.…”

“…Co-treatment of cells with that H 2 O 2 generation is likely associated with basal defense mechanisms. 10 In the present study, we further investigated the DHS signaling cascade leading to cell death in tobacco BY-2 cells, by evaluating the involvement of another key signaling molecule i.e., nitric oxide (NO). In plants, NO is known to play important roles in numerous physiological processes including germination, root growth, stomatal closing and adapative response to biotic and abiotic stresses (reviewed in ref.…”

“…Moreover, we recently showed in tobacco BY-2 cells that DHS triggers a dose-dependent production of H 2 O 2 via activation of a NADPH oxidase. 10 The DHS-induced cytosolic Ca 2+ transient is required for this H 2 O 2 production while the nuclear calcium variation is not necessary. In agreement with the results of Townley et al blocking the ROS production using diphenyleniodonium (DPI), a known inhibitor of NADPH oxidases, does not prevent DHS-induced cell death.…”

Maltose-binding protein switches programmed cell death in Nicotiana glutinosa leaf cells

“…In tobacco plants the new PCD inducers such as ultraviolet B overexposure [25] and dihydrosphingosine [26] have been reported, recently. We hope the present results may also open a new gate towards the studies with regard to the MBP-induced cell death in a plant cell and its signaling pathways in the future.…”

“…Plants also activate phenylpropanoid pathway and biosynthesis of antioxidant compounds during PCD induction [36]. However, a new report shown that dihydrosphingosine induces programmed cell death in tobacco cells independent of ROS [26].…”